PROJECT DESCRIPTION: Adult stem cells replenish cells lost due to normal attrition, and repair organs and tissues in response to injury. An excellent example of this phenomenon is the mammalian digestive epithelium, which completely renews its cells every 3-5 days under normal conditions, and can also replace cells lost to damage by radiation or cytotoxic substances (e.g. chemotherapy). This ability to rapidly self- renew is of great interest both because it predisposes the gastrointestinal system to cancer, and because the ability to promote renewal in specific disease contexts (e.g. short bowel syndrome) would be clinically beneficial. Additionally, an understanding of gastrointestinal repair may advance the more general application of stem cell technology to regenerative medicine. Despite recent progress, however, the molecular mechanisms regulating gastrointestinal stem cell maintenance, proliferation, and differentiation are not well understood. Because of their remarkable regenerative abilities and numerous recent advances in the application of modern molecular techniques, planariansare an attractive model organism for investigating tissue renewal and stem cell biology. In planarians, adult stem cells called neoblasts replenish tissues during normal cell turnover and during regeneration after injury. As one of the most prominent organs in the animal, the intestine of planarians offers an excellent opportunity to approach the molecular mechanisms linking injury and cell turnover to stem cell proliferation and lineage restriction. The experiments proposed capitalize on established techniques to identify genes expressed by planarian intestinal epithelial cells required for intestinal renewal and regeneration. The specific aims of this proposal are (1) to characterize the dynamics of tissue renewal and regeneration in the intestine by monitoring the differentiation of BrdU-labeled neoblasts;(2) to identify genes that are differentially expressed by intestinal epithelial cells by flow sorting these cells and conducting microarray analysis of gene expression;and (3) to identify genes that function autonomously and non-autonomously during neoblast proliferation and differentiation into intestinal epithelial cells in a small-scale, targeted RNA interference screen. PUBLIC HEALTH RELEVANCE: Adult stem cells normally replenish and repair the digestive tract throughout life. Deeper understanding of this phenomenon is required for effective treatment of digestive cancers and other diseases. In this proposal genes involved in this process will be identified in a model organism that can renew and completely regenerate its intestine.